Alcohol consumption, whether acute or chronic, results in increased susceptibility to infection and reduced antigen-specific immune activation. Monocytes and dendritic cells (DCs) of the innate immune system recognize pathogens and initiate antigen-specific T cell activation. Dendritic cells, depending on their subsets and maturation stage can augment or suppress T cell activation. Our preliminary results show that alcohol treatment results in reduced DC antigen presenting function which appears to be similar to tolerogenic dendritic cells described in the liver. We hypothesize that cells of the innate immune system are the primary targets of immunomodulation by alcohol. Specifically, alcohol inhibits accessory cell function of myeloid dendritic cells (DC) and alters innate immune recognition to result in impaired T cell activation. We postulate that several aspects of alcohol-induced T cell abnormalities are related to impaired dendritic cell functions either as a result of impaired accessory celI-T cell interactions or disturbed induction of cytokines. We further hypothesize that alcohol modulates TLR-mediated and alloantigen-induced cytokine induction in dendritic cells thereby affecting polarization of T cells. We propose that increased IL-10 and reduced IL-12 production in innate cells after acute alcohol use play a central role in inhibition of DC differentiation to result in an inhibitory DC phenotype that supports regulatory T cell development and activation. The specific aims of the proposal are: 1) To explore mechanisms that account for inhibition of myeloid DC differentiation by acute alcohol in vivo and in vitro by investigating a) intracellular pathways leading to increased IL-10 production in DCs, b) the autocrine role of alcohol-induced IL-10 and reduced IL-12 production on DC differentiation and maturation, c) intracellular pathways leading to reduced IL-12 production in DCs, d) the gene expression profile of alcohol-exposed early DCs. 2) To evaluate the role of DCs in inhibition of antigen-induced T cell responses after acute in vivo and in vitro alcohol treatment by testing a) the inhibitory potential of alcohol-exposed DCs on T cell activation by alloantigens b) induction of regulatory T cells (Tregs) determined by expression of CD4+CD25+, and c) changes in immunoinhibitory and stimulatory cytokine production profile triggered by various TLRs in mononuclear cells, monocytes and dendritic cell subsets. 3) To investigate the effects of acute alcohol intake and in vitro alcohol treatment on DC-mediated T cell polarization by evaluating a) Th1 and Th2 cytokine production profile in CD4+ T cells after co-culture with alcohol-treated DCs and b) transcriptional markers of Th1 (T-bet) and Th2 (GATA-3) subsets. These studies on myeloid DCs and their interactions with T lymphocytes should provide novel insights into the fine-tuning of alcohol-mediated immune regulation related to infections and to induction of autoimmune processes. Further, our studies will delineate some of the intracellular pathways that result in abnormal dendritic cell functions and contribute to aberrant antigen-specific T cell activation after alcohol use. Investigation of the modulation of IL-10 and IL-12 by alcohol in dendritic cell function may identify biomarkers of alcohol-induced immune depression and reveal potential targets for therapy.